Undesirable and dangerous side effects and adverse drug interactions are well known for the predominantly synthetic organic pharmaceuticals that have been widely administered over the past several decades. These adverse effects have led many research groups to go back and study, in greater detail, the medicinal properties and mechanisms of action of many natural compounds. Ancient cultures have long been aware of the medicinal properties of natural products, such as honey, compounds derived from botanical sources, and compounds from the seas. The subject matter of the present invention involves novel medicinal activities associated with natural products.
In one embodiment of the present invention, various antibacterial mechanisms are combined into a honey. Previously, different antibacterial mechanisms have been known to exist only separately in honeys derived from different floral sources. Honey has been widely accepted as both food and medicine by most, if not all, generations, traditions, and civilizations, both ancient and modern. Although honey has been used by humans for more than 5,000 years to treat a variety of ailments, it has been recognized for almost as long that honeys derived from some floral sources are more medicinal than others. As a general rule, darker honeys have more medicinal activities than light honeys. Many studies have shown that medicinal honey influences biological systems as antioxidant, anti-inflammatory, and antimicrobial. In addition, honey acts as an autolytic debridement agent on wounds, as a cough suppressant, analgesic, remedy for dyspepsia, and natural anti-cancer agent.
One of the darkest honeys is buckwheat honey, which has been shown to have one of the highest antioxidant, anti-inflammatory, and antibacterial activities of any honey variety tested. Because of the bacterial resistance problems that have arisen from the overuse and misuse of antibiotics, the antibacterial activity of honey is the activity that has renewed the interest in honey, particularly for treatment of hard-to-heal (chronic) wounds. But the antibacterial activity of honeys derived from different floral sources has been found to be due to different mechanisms. Early on, honey's antibacterial activity was attributed to its osmotic effect and to its low pH, but these have subsequently been found to contribute only minor antibacterial effects. The first factor discovered that contributes a major antibacterial activity in honey was hydrogen peroxide, but its generation and concentration are under the control of a number of important effects.
First, hydrogen peroxide is not a constituent of the nectar from which honey is produced. It is derived from the enzymatic activity of glucose oxidase acting on glucose. The maturation of honey from plant nectar is dependent upon the activities of several enzymes, most of which are derived from the hypopharyngeal gland of the honey bee. Diastase (amylase), derived from the bee, breaks down starch to smaller carbohydrates (dextrins, oligo-, di- and monosaccharides [glucose]). Invertase, derived from the bee, converts sucrose, the primary sugar in nectar, into glucose and fructose. Glucose oxidase, also derived from the bee, catalyzes the oxidation of glucose by molecular oxygen to gluconolactone, which subsequently hydrolyses spontaneously to gluconic acid and hydrogen peroxide. Gluconic acid is the primary acid in honey responsible for most of honey's acidity and low pH, and hydrogen peroxide is the primary antibacterial agent in most medicinal honeys.
Second, the production of hydrogen peroxide is very slow in mature honey for two reasons: i) the activity of glucose oxidase is depressed by high osmotic pressure, and ii) the spontaneous conversion of gluconolactone to glucuronic acid and hydrogen peroxide is a hydrolysis reaction requiring water, which is unavailable in ripe honey. Most hydrogen peroxide present in ripe honey is generated as the honey is being ripened and dried by the bees. And when ripened honey is subsequently diluted, by wound fluid for example, this reaction speeds up again. Upon dilution of medicinal honey, the rate of hydrogen peroxide generation is continuous and can reach concentrations up to 4 mmol/L, with a mean of about 1-2 mmol/L. This relatively low concentration is nevertheless high enough to provide a substantial antibacterial activity, and yet is about 1000-times less than the 3% solution commonly used as an antiseptic; which has been associated with tissue damage, including damage to fibroblast cells from human skin. Furthermore, the continuous production of hydrogen peroxide in diluted honey produces a long-lasting antiseptic effect that is most sought after in fighting infections in wounds. It has been reported that hydrogen peroxide is more effective when supplied by continuous generation from glucose oxidase catalysis, as in honey, than when added as a single bolus.
Third, in addition to the glucose/glucose oxidase system as a main source of hydrogen peroxide generation, plant-derived polyphenols present in some honeys provide a supplementary source of hydrogen peroxide. Honeys with high concentrations of polyphenols, such as buckwheat honey, have higher hydrogen peroxide levels due to this second method of hydrogen peroxide generation. The mechanism of this action is likely from the auto-oxidation of polyphenols yielding both hydrogen peroxide and phenoxyl-radicals. Furthermore, redox-active phenolics appear to be active intermediates that confer additional oxidative activity on hydrogen peroxide. In addition, the chemical interaction of honey phenolics with hydrogen peroxide results in products that degrade bacterial DNA. In the presence of transition metal ions, via the Fenton reaction, hydrogen peroxide is also converted to hydroxyl radicals. Both the phenoxyl- and hydroxyl-radicals have been shown to induce strand breaks in DNA. Thus, a second factor present in some honeys that contribute to its antibacterial effect are polyphenols.
A third factor found in honey that has antibacterial activity is methylglyoxal (MGO), but this agent has only been found in honey derived from certain floral species of the Leptospermum genus of shrubs and small trees found in New Zealand, Australia, Malaysia, and Indonesia. Originally referred to as UMF (Unique Manuka Factor), methylglyoxal has been found to originate in honey from dihydroxyacetone present in the nectar of Leptospermum flowers, for example from the manuka tea tree (Leptospermum scoparium) of New Zealand or the jelly bush (Leptospermum polygalifolium) of Australia. Since the first description of UMF, it has been recognized that its concentration is highly variable in different manuka honey batches, and that has been determined to be due to different concentrations of dihydroxyacetone in different cultivars of manuka, with pink-flowered cultivars producing the highest dihydroxyacetone levels in nectar. There are also seasonal changes within a Leptospermum species, or between the different species. Because of this batch-to-batch variability, the methylglyoxal levels or antibacterial activity of each lot of Leptospermum honey must be assayed to determine whether it will be useful as a medicinal honey or not. As manuka honey often has very low levels of hydrogen peroxide, methylglyoxal becomes its primary antibacterial agent.
A fourth antibacterial factor that has been found in Revamil Source honey that is produced in greenhouses in The Netherlands is Bee Defensin-1, a cationic antimicrobial peptide placed in this honey variety by the bees. Defensins are antimicrobial peptides found in many organisms, including plants, invertebrates, insects, birds and mammals. They are cysteine-rich peptides with multiple disulfide bonds and a triple-stranded beta sheet. Most defensins function by binding to the microbial cell membrane, and once embedded, they form pore-like membrane defects that allow efflux of essential ions and nutrients. Bee Defensin-1, a 51-amino acid peptide (also called Royalisin because it was first discovered in royal jelly), was discovered in Revamil Source honey when bactericidal activity was not eliminated by neutralization of the usual antimicrobial factors (hydrogen peroxide and methylglyoxal). The activity was found in a relatively high molecular weight (>5-kDa) chromatographic fraction; stained as a protein on polyacrylamide gel electrophoresis, and was immuno-stained by anti-bee defensin-1 antibody on a Western blot. In addition, the antibacterial activity of Revamil Source honey was abolished by proteolytic digestion with pepsin and by the anti-bee defensin-1 antibody.
Medicinal honeys from different floral sources exhibit differing antibacterial activities towards different bacterial pathogens. For example, Mundo et al., (2004) reported varying sensitivities to the antibacterial properties of 26 different honey types by nine different bacteria, including multiple strains of Staphylococcus aureus, emphasizing the variability in the antibacterial effect of different honey samples. These authors reported that whereas Bacillus stearothermophilus was the most sensitive microorganism to the antibacterial activity of medicinal honeys in this study, Alcaligenes faecalis, Lactobacillus acidophilus, and Staphylococcus aureus strains ATCC 25923, 8095, and 9144 were each moderately sensitive, and Escherichia coli, Salmonella enterica, Pseudomonas fluorescens, Bacillus cereus, and Listeria monocytogenes were the most resistant to the antibacterial activity of honey.
In this study it was demonstrated that different microorganisms were more or less susceptible to the different antibacterial mechanisms in various honeys. Whereas it required 50% manuka honey with its non-peroxide methylglyoxal antibacterial mechanism to inhibit the growth of B. stearothermophilus, buckwheat honey at only 25% concentration was required to inhibit the growth of this organism via its hydrogen peroxide-dependent antibacterial action. The same was true for the inhibition of S. aureus strains ATCC 9144 and 25923 which both were inhibited by 50% manuka honey but by only 33% buckwheat honey, whereas the converse was true for the inhibition of S. aureus strain ATCC 8095 and B. cereus where 50% buckwheat honey was required to completely inhibit their growth while only 25% manuka honey was required. Table 1 summarizes the bacterial sensitivities of the various bacteria to the different honeys.
TABLE 1Bacterial Sensitivity by Type andInhibitory Concentration of Honey.Type of Honey and (Inhibitory Concentration;BacteriaBacteria % honey in water, w/v)E. coli O157:H7christmas berry (100); saw palmetto (100);tarweed (100); buckwheat (100); manuka (50)S. entericamanuka (50)A. faecalisblueberry (100); soybean (100); tarweed (33);buckwheat (33); manuka (25); horsemint (25)P. fluorescenstarweed (100); buckwheat (50)L. acidophilussoybean (100); christmas berry (100);buckwheat (100); manuka (100); saw palmetto(100); melaleuca (50); tarweed (50)L. monocytogenesmelaleuca (100); tarweed (100); buckwheat(100)B. cereustarweed (100); buckwheat (50); manuka (25)S. aureus ATCCchristmas berry (100); saw palmetto (50);8095tarweed (50); buckwheat (50); cotton (33);manuka (25)S. aureus ATCCsaw palmetto (100); sunflower (100); horsemint9144(100); manuka (50); melaleuca (33); buckwheat(33)S. aureus ATCCsoybean (100); sunflower (100); saw palmetto25923(50); melaleuca (50); rabbit bush (50); manuka(50); tarweed (33); buckwheat (33)B. Stearothermophilusblueberry (100); blackberry (100); manuka (50);black sage (50); red sumac (50); melaleuca (50);horsemint (50); christmas berry (50); soybean(33); alfalfa (33); cotton (33); saw palmetto(33); rabbit bush (33); tarweed (25); buckwheat(25); knotweed (20); sunflower (17)
Data from Mundo et al., 2004.
Of the honeys listed in Table 1, buckwheat, tarweed, saw palmetto and melaleuca inhibit bacteria primarily via hydrogen peroxide, whereas the antibacterial activity of manuka, blueberry, and knotweed honeys is primarily non-peroxide mediated. Other studies report similar findings and therefore the present disclosure relates to a honey composition containing high concentrations of both peroxide and non-peroxide antibacterial activities in order to produce a honey with broad-spectrum antibacterial activity efficient at inhibiting growth of most major wound pathogenic bacteria at one low honey concentration.
Plants have also been used for medicinal purposes since before recorded history. Ancient Chinese and Egyptian papyrus writings describe medicinal uses for plants as early as 3,000 BC. While some cultures, such as Africans and Native Americans, have used botanical sources in their healing remedies, other cultures including the Chinese and Indians have developed medicinal systems, such as Traditional Chinese Medicine and Ayurveda, respectively, in which botanicals were used. Modern researchers have found that people in different parts of the world used the same or similar plants for the same purposes.
In the early 19th Century, when chemical analysis first became available, scientists began to extract and modify the active ingredients from botanical sources. Today almost one quarter of pharmaceutical drugs are derived from botanicals. Recently, the World Health Organization (WHO) estimated that 80% of people worldwide rely on botanical medicines for some part of their primary health care. Botanical medicine is being taught more in medical schools and pharmacy schools, and more health care providers are learning about the positive effects of using botanical medicine to help treat health conditions. In Germany, for example, about 600-700 plant-based medicines are available and are prescribed by some 70% of German physicians. In the past 20 years in the United States, public dissatisfaction with the cost of prescription medications and their extensive adverse effects, combined with an interest in returning to more natural remedies, has led to an increase in botanical medicine use. Botanical medicine is used to treat many conditions, such as asthma, eczema, premenstrual syndrome, menopausal symptoms, rheumatoid arthritis, migraine, chronic fatigue, irritable bowel syndrome, cancer and chronic wounds among others. Today, nearly one-third of Americans use herbs and one study found that 90% of arthritic patients use alternative therapies such as botanical medicine.
One such botanical remedy originated in American Indian folklore as a treatment for hard-to-heal wounds. It was handed down to the pioneers in South Carolina as an extract of ash derived specifically from Red Oak Bark grown in the region around Piedmont, S.C. A paste of this extract that was applied to a hard-to-heal wound or skin ulcer proved to be very effective at healing the wound. It is now known that Red Oak Bark contains specific storage cells that collect and concentrate certain mineral ions from the soil in which they grow, and that the soil around Piedmont, S.C. contained a high level of specific minerals that are key for healing wounds. Chemical analyses of the extracts from the ash of Red Oak Bark led to the identification of the minerals responsible for the wound healing effect and a safe and efficacious version of the active minerals, which included potassium, calcium, zinc and rubidium is now manufactured by combining these minerals in a proprietary formulation and used for chronic wound therapy. U.S. Pat. No. 5,080,900 described the oak bark ash extract and U.S. Pat. Nos. 6,149,947 and 7,014,870 described the synthetic version of the formulation. All three patents are now expired.
In one study, Weindorf et al., (2012) demonstrated that the synthetic formulation of these ions prepared in the pharmaceutically-accepted carrier of polyethylene glycols used to treat over 300 therapy-refractory wounds, demonstrated a wound size reduction of at least 50% in 73% of the patients, where a wound size reduction of at least 50% is predictive of successful wound closure.
As with plants, the marine ecosystem has been a source of therapeutics for the treatment of human diseases. In recent times, cancer drugs have been developed from marine sources. Cytosar, a staple treatment for leukemia and lymphoma was derived from a Caribbean sea sponge. Other anti-cancer drugs have been derived from tunicates and a potent medicine for the treatment of chronic pain, more powerful than morphine, has been derived from the venom of cone snails that inhabit the reefs of Australia, Indonesia and the Philippines. The waters of the Dead Sea have been renowned for their therapeutic effects since ancient times. Galenus, a prominent first Century Greek physician, stated that this salt water was good for the treatment of arthritis, eczema, muscular pain, rheumatism, and psoriasis, and the Jewish-Roman historian, Flavius Josephus, wrote two thousand years ago that the salts from the Dead Sea heal the human body and are therefore used in many medicines.
The water of the Dead Sea is unique compared to other seas and lakes in its high concentration of salts--Dead Sea water contains 330 g of minerals per liter (33%). This salt concentration is between 7-10 times that of the oceans, which typically contain 3.5% minerals. The mineral composition of the Dead Sea is also significantly different from that of ocean water. Whereas the major salt constituent of ordinary seawater is sodium chloride (NaCl), Dead Sea salt is rich in MgCl.sub.2, CaCl.sub.2, KCl, MgBr.sub.2 and CaSO.sub.4. The concentration of ionic species present in the Dead Sea water is: magnesium (40.65 g/L), sodium (39.15 g/L), calcium (16.86 g/L), potassium (7.26 g/L), chloride (212.4 g/L), bromide (5.12 g/L), sulfate (0.47 g/L), and bicarbonate (0.22 g/L). The bromide ion concentration is the highest of all waters on the earth and serum bromide levels have been shown to increase up to 4-fold after bathing in the Dead Sea for four weeks, as a result of entering the circulation and internal organs through the skin. Metal ions are required for many critical functions in humans. Scarcity of some metal ions often leads to disease. Four main group metals (Na, K, Mg, and Ca) and 10 transition metals (V, Cr, Mn, Fe, Co, Ni, Cu, Zn, Mo, and Cd) are currently known or thought to be required for normal biological function. It is believed that the therapeutic properties of the Dead Sea are due to a large extent to the presence of magnesium, potassium and bromide.
Various cultures and groups of people have visited the Dead Sea for therapy, dating back to the time of the ancient Egyptians, utilizing the salt in various unguents and skin creams, as well as soaps, just as it is used today. The Dead Sea has taken on a new dimension today: modern science has proven the therapeutic and rejuvenating properties of its unique mineral content. The Dead Sea has become a renowned center for natural health with people coming from around the world to bath in its mineral-rich waters. Balneotherapy emerged as an important treatment modality in the 1800s, first in Europe and then in the United States. After seeing decline in use for almost 50 years, at about the same time as the decline in the use of medicinal honey, balneotherapy in the Dead Sea has experienced resurgence in popularity over the past two decades at the same time as a new recognition of the safety and efficacy of natural remedies has fueled a resurgence in popularity for these treatment modalities. The major dermatological diseases that are frequently treated by baloneotherapy in the Dead Sea with a high rate of success are psoriasis and atopic dermatitis. Both magnesium and potassium ions from the Dead Sea have a specific inhibitory capacity on the uncontrolled proliferation of psoriatic dermis grown in tissue culture.
Magnesium salts, the prevalent minerals in Dead Sea salt, are known to exhibit favorable effects in inflammatory diseases. In one study of atopic dry skin, bathing in magnesium-rich Dead Sea salt solution improved skin barrier function, enhanced skin hydration, and reduced inflammation. In other in vitro and in vivo studies, magnesium ions inhibit the number and function of epidermal Langerhans cells that contribute to inflammatory skin diseases by presenting alloantigens to T lymphocytes, thereby activating these cells to release pro-inflammatory cytokines. In one study, the reduced antigen presenting activity of MgCl.sub.2-treated Langerhan's cells was associated with suppression of constitutive tumor necrosis factor (TNF)-.alpha. production by the epidermal cells in vitro. Another study showed that Dead Sea water inhibited the production and/or release of the pro-inflammatory cytokines, interleukin (IL)-2 and interferon (IFN)-.gamma., from Th1 lymphocytes. Others confirmed down-regulation of the pro-inflammatory cytokines, TNF-.alpha. and IL-1, and an up-regulation of insulin-like growth factor (IGF)-1 following balneotherapy in the Dead Sea.
Magnesium impacts more than 325 enzyme systems in the human body. For example, it is a rate-limiting factor in the activation of epidermal adenylate cyclase and consequently in the production of cyclic adenosine monophosphate (cAMP). A decrease of cAMP and concomitant increase of cyclic guanosine monophosphate (cGMP) has been implicated in excessive cellular proliferation, a major element of the psoriatic state. Balneotherapy in Dead Sea water has also been applied to the treatment of various inflammatory rheumatic diseases such as rheumatoid arthritis and psoriatic arthritis. In a study of Dead Sea water in the treatment of patients with rheumatoid arthritis, Dead Sea water produced statistically significant clinical improvements in most parameters for up to one month following cessation of treatment, whereas treatment with sodium chloride water did not. Dead Sea balneotherapy for 14 days also produced significant clinical improvements in knee osteoarthritis, which lasted for at least 1 month following cessation of treatment. Many other diseases are also treated by balneotherapy in the Dead Sea, including chronic ulcers.
Hypertonic Dead Sea water has also been shown to be efficacious in the treatment of allergic rhinitis. The two common treatments for such sinonasal disease are intranasal rinsing with normal saline solution or corticosteroid rinses, and both have positive effects on the physiology of nasal mucosa. In a study comparing Dead Sea saline spray with NaCl solution, the hypertonic Dead Sea saline solution proved efficacious in mild-to-moderate allergic rhinitis, including improving mucociliary clearance, whereas no significant improvement was seen with the nasal saline spray.
DePootere et al., (2011) demonstrated that the addition of magnesium and bromide to some of the botanically-derived inorganic metal ions gave an enhanced anti-inflammatory and wound healing effect in an animal model of chronic rhinosinusitis. The present invention adds antibacterial and anti-biofilm activities to the anti-inflammatory activity of the mineral salts composed of a mixture of active ingredients from two sources: a tree bark, and the Dead Sea.
As can be derived from the variety of devices and methods directed at wound healing compositions, many strategies have been contemplated to accomplish the desired end. Heretofore, widely administered synthetic organic pharmaceuticals are commonly associated with undesirable side effects and adverse drug interactions. Thus, there is a long-felt need for more natural wound healing compositions. There is a further long-felt need for wound healing compositions involving medicinal honey, mineral ions, and methylglyoxal, and their corresponding methods of use.